The ability to use radiolabeled peptides or peptidomimetics as diagnostic or therapeutic medical tools has long been a goal of the pharmaceutical industry. This goal is being realized as radiolabeled peptide products have now been introduced into the marketplace. For example, an Indium-111 labeled pentatreotide product has been introduced for somatostatin receptor imaging for neuroendocrine tumors (Mallincrodt Medical, Inc.). As this type of product line matures it will become increasingly important to be able to prepare the radiolabeled peptide product in an efficient, cost-effective manner and in high purity.
The labelling of the peptide with the radionuclide has been one of the primary difficulties in bringing a radiolabeled peptide to the market. It is important that the radionuclide be stably coupled to the peptide when administered in vivo while also not interfering with the biological activity of the peptide. Labelling the peptide directly with a radionuclide is not acceptable because of the possibility that the radionuclide could attach at an important biological site on the peptide and interfere with its activity or specificity. As an alternative to direct labelling, the use of a "bifunctional chelate" in which a composition having a strong chelating group capable of covalently bonding to the peptide and being labelled with the radionuclide has been proposed. One such approach is described in U.S. Pat. No. 4,479,930 which issued to D. J. Hnatowich. In the Hnatowich patent, the method for radiolabelling peptides involves the reaction of a peptide with a dicyclic anhydride of a defined formula (generally a DTPA-type or EDTA-type compound) and then adding a radionuclide to the peptide/chelate conjugate. Even though useful in the radiolabelling of a peptide, the method described in Hnatowich has one significant drawback; it is not capable of specifically coupling only one peptide per chelate and, in practice, the resulting peptide/chelate conjugate is predominantly formed as a diaddition product. The diaddition product is not clinically useful and must be removed by lengthy, complicated and costly purification processes. Furthermore, valuable peptide is lost in this process. This problem is further illustrated in Example 1 of PCT International Publication Number WO 90/06949.
Most prior attempts at radiolabelling peptides have used standard organic chemistry procedures to couple the chelating group to a previously prepared peptide. It would be advantageous if the chelating group could be incorporated into the peptide as the peptide is being formed using solid phase peptide synthetic chemistry. Kazmierski [Tet. Letters, 4493 (1993); and Int. J. Peptide and Protein Res., 45, 241-247 (1995)] was concerned with synthesizing metal binding peptides incorporating aminodiacetic acid based ligands, but suggested that prederivitized amino acids containing the ligand be prepared and the peptides incorporated during peptide synthesis. Moreover, Kazmierski requires that one of the bonding atoms for the metal come from the nitrogen group of the amino acid which could adversely affect the biological activity of the peptide. This significantly limits the usefulness of this approach. Sluka, et al. [J. Amer. Chem. Soc., 112, 6369 (1990)] described a method for the solid-phase synthesis of protein-EDTA for use in affinity cleaving, but this method was not adapted for use with polyaminocarboxylic acids nor for binding the medically useful lanthanide and actinide radionuclides. W. B. Edwards, et al. [J. Med. Chem., 37, 3749 (1994)] discolsed the coupling of DTPA-bis-anhydride to the N-terminal D-Phe of the heptapeptide attached to HMP-resin followed by aminolysis to generate the mono DTPA derivative in 5% yield. The low yield can be attributed to formation of the diaddition DTPA-(peptide).sub.2 derivative.
There is, therefore, a need for a method for radiolabelling peptides utilizing polyaminocarboxylate ligands of the DTPA or EDTA type in a solid phase peptide synthesis process in which no undesirable diaddition products are formed and only the desired monoaddition product is formed in high purity.